Project Summary microRNAs (miRNAs) play critical regulatory roles in multiple types of skeletal cells. Despite the accumulating evidence for miRNAs' involvement in multiple processes of skeletal development and homeostasis, specific roles of individual miRNAs in vivo are largely unknown. miR-17-92 miRNAs are abundantly expressed in stem/progenitor cells and are implicated in their maintenance by promoting proliferation and suppressing differentiation. miR-17-92 cluster miRNAs are so far the only miRNAs whose genetic mutations have been demonstrated to cause human congenital skeletal diseases; duplications of the miR-17-92 gene cause macrocephaly and digit abnormalities, whereas its heterozygous deletions result in Feingold syndrome II that is characterized by macrocephaly and brachysyndactyly. In addition, we have also found that miR-17- 92 miRNAs negatively regulate osteoblast differentiation and bone mass in mice. However, the mechanisms by which miR-17-92 miRNAs regulate skeletal development and bone homeostasis are unknown. Because miR-17-92 miRNAs are predicted to target more than 2,000 genes, identifying critical molecular mechanisms is a challenge. On the other hand, it is presumed that physiologically important miRNA-RNA interactions are evolutionarily selected to control certain biological processes and pathways, as miR-17-92 miRNAs have been reported to regulate several signaling pathways in a cellular context dependent manner. Based on these findings, we hypothesize that miR-17-92 miRNAs regulate key signaling pathways in skeletal stem/progenitor cells to control their proliferation and differentiation, and thus regulate skeletal development and bone homeostasis. To test this hypothesis, we will first generate mouse models to define the role of miR-17-92 miRNAs in skeletal development and postnatal bone homeostasis at the tissue and cellular levels in vivo. We will then dissect molecular mechanisms that mediate miR-17-92 miRNAs' action at the levels of miRNA-RNA interaction, gene expression, and signaling pathways. This project will define the role and mechanisms of action of miR-17-92 miRNAs in skeletal stem/progenitor cells during skeletal development and bone homeostasis and will provide the basis for developing new therapeutic strategies for miR-17-92-related diseases and for common skeletal diseases such as osteoporosis.